Two kinds of peptide hormones, named "atrial natriuretic peptide (ANP)" and "brain natriuretic peptide (BNP)", have recently been isolated from mammalian atria and brains as hormones that regulate the homeostatic balance of body fluid volume and blood pressure. The structures of those peptides and the mechanism of their biosynthesis have been unravelled and their physiological actions are also being unravelled.
ANP was first isolated from the human atrium in three types, .alpha.-type having a molecular weight of ca. 3000 (.alpha.-hANP), .beta.-type of ca. 6000 (.beta.-hANP) and .gamma.-type of 13000 (.gamma.-hANP), and their respective structures were unravelled (Kangawa, K. et al., Blochem. Biophys. Res. Commun., 118, 131, 1984; Kangawa, K. et al., Nature, 313, 397, 1985).
As a result, the following facts have been determined: (1) .alpha.-hANP is a single-stranded peptide that consists of 28 amino acids having a single S-S bond in the molecule; (2) .beta.-hANP is an antiparallel dimer having an S-S bond formed between the molecules of .gamma.-hANP; and (3) .gamma.-hANP is a high-molecular weight protein composed of 126 amino acids, with .alpha.-hANP being contained in the C-terminal portion.
Further, analysis of the cDNA coding for .alpha.-hANP has shown that each of those three types of hANP (.alpha.-, .beta.- and .gamma.-hANP) is biosynthesized from the same precursor protein (Oikawa, S. et al., Nature, 309, 724, 1984). Stated more specifically, those peptides are first biosynthesized in atrial cells as a precursor (pre-hANP) composed of 151 amino acid residues and then, the signal peptide composed of 25 N-terminal residues is cleaved in the Golgi body to produce .gamma.-hANP. Subsequently, the .gamma.-hANP is further cleaved with an enzyme (i.e., subjected to processing) for transformation to .alpha.-hANP, which is secreted primarily into blood. The process of .beta.-hANP synthesis still remains unclear today but most probably it is produced by way of .alpha.-hANP.
Ever since the structure of hANP was first unravelled, the structures of ANPs derived from other mammals have also been studied. And, today, the following knowledge is available: ANPs have similar amino acid sequences over a broad spectrum of mammals ranging from rodents to humans; in particular, .alpha.-type ANP (.alpha.-ANP) has the same amino acid sequence in higher mammals including humans, dogs and pigs; and .alpha.-type ANPs derived from rats and rabbits have entirely the same amino acid sequence as .alpha.-hANP except that the methionine residue in position 12 is replaced by an isoleucine residue (0ikawa, S. et al., Blochem, Biophys. Res. Commun., 132, 892, 1985; Forssmann, W.G. et al., Anat. Embryol., 168, 307, 1983).
When the distribution of ANP in vivo was examined using anti-.alpha.-hANP antisera, it was found that ANP also occurred in the brain, though in small amounts, as well as in the atrium. Further, ANP-containing neurons have been reported to occur in the hypothalamus and pontine tegmentum of the brain (Cantin, M. et al., Itistochemistry, 80, 113, 1984; Saper, C.B., et al., Science, 227, 1047, 1985) and, therefore, it is speculated today that ANP may also work in the brain as a nerve transmitter that participates in the regulation of the cardiovascular system.
Recently, a new peptide that was similar in structure to ANP but that was clearly distinguishable from the latter was isolated and identified from porcine brain and, like ANP, this peptide was verified to have natriuretic and hypotensive actions and hence was named "BNP" (Sudoh, T. et al., Nature, 332, 78, 1988). It was later found that porcine-derived BNP (pBNP) was a single-stranded peptide composed of 26 amino acids having a single S--S bond in the molecule. Further, a cDNA coding for human BNP was isolated and the structure of tile BNP precursor also became clear, showing that BNP was constructed from an entirely different precursor than in the case of ANP (Sudoh, T. al al., Biochem. Biophys. Res. Commun., 159, 1427, 1989). As of today, the structure of rat BNP has also been unravelled (Kojima, M. et al., Biochem. Biophys. Res. Commun., 159, 1420, 1989).
It was also found that BNP was present in porcine brain in an amount ten times higher than ANP and this suggested a higher possibility that in the brain BNP would work as a nerve transmitter for the nerve system to regulate the homeostatic balance of body fluid volume and blood pressure (Ueda, S. et al., Blochem. Biophys. Res. Commun., 155, 733, 1988).
It was later found that like ANP, BNP occurred not only in the brain but also in the atrium (though in an amount of only 2- 3% of ANP) to be secreted into blood, showing that like ANP, BNP was also a hormone regulating the homeostatic balance of body fluid volume and blood pressure (Minamino, N. et al., Blochem. Biophys. Res. Commun., 155, 740, 1988: Aburaya, M. et al., Biochem. Biophys, Res. Commun., 165, 872, 1989). As a matter of fact, it has been verified with experimentation on rats that pBNP has comparable levels of natriuretic and hypotensive actions to .alpha.-hANP. Thus, it has been found to date that at least two obviously different types (ANP and BNP) of hormones occur in mammals and that they regulate the homeostatic balance of body fluid volume and blood pressure. These peptides are secreted from the atrium into blood and work as hormones that regulate the homeostatic balance of body fluid volume and blood pressure. Further, it has been found that those peptides also occur in the brain, where they work as nerve transmitters for the nerve system to regulate the homeostatic balance of body fluid volume and blood pressure. At the same time, the studies conducted to date have shown that three kinds of receptor eDNA are cloned to those peptides and their structures have been identified. Two of the three receptor types have a guanylate cyclase domain in a intracellular portion of the molecule and the other type, which is generally referred to as a C-receptor (clearance receptor), does not have a guanylate cyclase domain in a intracellular portion of the molecule (Chinkers, M. et al., Nature, 338, 78, 1989; Chang, M.S. et al., Nature, 341, 68, 1989; Schulz et al., Cell, 58, 1155, 1989; Fuller, F. et al., J. Biol. Chem., 263, 9395, 1988).
However, as of today, the relationships between those receptors and the ligands (ANP, BNP) have not been clearly established. In other words, much still remains unclear about the relationships between the individual receptors and the physiological actions developed, as well as the specificity between ligand and receptor.
With all points of the foregoing discussion taken into account, the question arises as to whether ANP and BNP known today are the only hormones in mammals that regulate the homeostatic balance of body fluid volume and blood pressure. In particular, considering the aforementioned versatility of receptors, there is a high possibility that a new third peptide hormone (novel ligand) might exist aside from ANP and BNP.
However, as of today, whether any such novel ligand exists remains unclear.
Therefore, the object of the present invention is to isolate from mammals a novel peptide hormone that exhibits physiological activities (e.g. natriuretic and hypotensive actions) similar to ANP and BNP already known in mammals but which is clearly distinguishable from them and to establish a method of presenting said peptide to the industry.